Gram-negative bacterial diseases are important causes of morbidity and mortality throughout the world. Our research will examine three components of the interaction between Gram-negative bacteria and the host, focusing on the role of Gram-negative bacterial lipopolysaccharides (LPS; endotoxin) in pathogenesis. First, we will characterize the LPS which are released by Gram-negative cells growing in vitro and in vivo, as well as the LPS shed by cells exposed to plasma. Second, we will analyze the associations between LPS and human lipoproteins, in order to understand the role of lipoproteins in modifying the toxicity of LPS in plasma and the uptake of LPS by cells. Third, we will correlate the uptake and degradation of LPS by human monocytes and polymorphonuclear leukocytes with the release of mediators (prostaglandins) by these cells. Sensitive immunoassays will be developed to measure the soluble products of LPS degradation. Our experiments will differ from previous work by (1) using defined bacterial mutants which allow intrinsic radiolabeling of LPS, protein, and phospholipid, in order to study the composition of LPS-containing material shed from bacterial cells (2) emphasizing the physical differences between purified LPS and the LPS which is present in bacterial membranes or shed from the bacterial surface. We hypothesize that these physical differences may influence the interactions of LPS with plasma and cells. These studies should lead to progress in (1) immunodiagnosis of Gram-negative infections, and (2) understanding the factors which influence LPS "processing" by the host, thus improving the scientific basis for advances in the therapy of diseases caused by Gram-negative bacteria.